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STUDIES OF SIR2 AND CALORIC RESTRICTION
MIMETIC PATHWAYS IN AGING
by
Jia Hu
A Dissertation Presented to the
FACULTY OF THE USC GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(MOLECULAR BIOLOGY)
May 2013
Copyright 2013 Jia Hu

Increased dosage of Sir2, a conserved histone deacetylase, extends replicative life span in yeast, and possibly worms, and flies and protects mammals against certain diseases. Previous work in our lab has shown that it is the lack of Sir2 and not its overexpression that promotes resistance to different stresses and extends chronological lifespan when combined with calorie restriction (CR) and/or mutations in the Tor/Sch9 or Ras pathways in yeast. To identify genes and pathways that mimic the effects of CR on aging and cellular protection we explored the role of Sir2 in regulating stress response and examined the genetic interaction between Sir2 and stress resistance transcription factors, Msn2/4 and Gis1. Our results suggest that Msn2/4 and Gis1 are required for the enhanced stress resistance of sir2Δ mutant and the further extension of chronological lifespan of sir2Δ mutant under caloric restriction. In agreement with this result, the serine/theronine kinase Rim15, a positive regulator of Msn2/4 and Gis1, was implicated in Sir2 mediated cellular sensitization to stress. We also examined the role of Sir2 in genomic stability during chronological aging and proposed a potential mechanism for its action. Further study of Sir2 implicated its involvement in regulating Ras2 expression. These studies shed light on the investigation of Sir2's function in higher eukaryotes. ❧ The Sir2 homolog SIRT1 deacetylase, one of the best-characterized sirtuins in mammals, has been shown to mediate some of the beneficial effects of calorie restriction (CR). However, its role in CR-dependent lifespan extension still remains unknown and highly controversial. We previously found that mice lacking both copies of SIRT1 displayed a shorter median lifespan than wild type mice on an ad libitum or a caloric resitriction diet. Here we demonstrated that the median lifespan of SIRT1+/- heterozygote mice in CR was identical to that observed in wild type mice but a higher frequency of pathologies was displayed in SIRT1+/- mice. Microarray gene expression analysis further revealed the possible relations between SIRT1 and CR. Our results suggest that some SIRT1 expression but not its high expression is required for the beneficial effect of CR in longevity and health. ❧ In mammals, several days of food deprivation lead to the accumulation of ketone bodies including acetoacetic acid in the blood. Here we show that as external glucose becomes depleted, S. cerevisiae convert leucine to the ketone body-like acetic acid, analogously to the conversion of leucine to acetoacetate in fasting mammals. Acetic acid promoted the activation of pro-aging pathways similarly to glucose and ethanol. Whereas wild type and ras2 mutant cells accumulated acetic acid, tor1 and sch9 mutants depleted it rapidly by a mechanism that required acetate CoA-transferase and that was essential for lifespan extension. In sch9mutants acetic acid was partly depleted by a mechanism that required oxidative phosphorylation and was utilized to promote the stress resistance carbon source trehalose generation. These results indicate that that the effect of Tor/S6K deficiency on CLS extension involves an alternative metabolic mode, in which acetic acid can be utilized for energy production and the storage of stress resistance carbon sources. These effects are reminiscent of those described for mammals in response to fasting and raise the possibility that the life span extending effect of inhibition of TOR/S6K in higher eukaryotes may also involve analogous metabolic switches.

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STUDIES OF SIR2 AND CALORIC RESTRICTION
MIMETIC PATHWAYS IN AGING
by
Jia Hu
A Dissertation Presented to the
FACULTY OF THE USC GRADUATE SCHOOL
UNIVERSITY OF SOUTHERN CALIFORNIA
In Partial Fulfillment of the
Requirements for the Degree
DOCTOR OF PHILOSOPHY
(MOLECULAR BIOLOGY)
May 2013
Copyright 2013 Jia Hu